本文關(guān)鍵詞： 軟骨組織工程 脫細(xì)胞真皮基質(zhì) 支架材料 Ⅱ型膠原　出處：《中國修復(fù)重建外科雜志》2014年06期 　論文類型：期刊論文

【摘要】：目的制備脫細(xì)胞真皮基質(zhì),檢測其細(xì)胞及組織相容性,探討其作為軟骨組織工程支架材料的可行性。方法取新生小牛背部真皮組織,以0.5%SDS溶液水平振蕩脫細(xì)胞,0.5%胰蛋白酶行膠原纖維結(jié)構(gòu)重塑,甲醛浸泡進(jìn)行交聯(lián),0.5%硫酸軟骨素溶液中超聲振蕩進(jìn)行表面修飾,制備脫細(xì)胞真皮基質(zhì)支架材料。乙醇排除法檢測結(jié)構(gòu)重塑前后材料孔隙率變化;MTT法檢測材料細(xì)胞毒性;將材料植入成年SD大鼠背部皮下,評價組織相容性。采用以3∶7比例共培養(yǎng)的第2代新西蘭大白兔軟骨細(xì)胞與骨髓基質(zhì)細(xì)胞作為種子細(xì)胞,種植于脫細(xì)胞真皮基質(zhì)支架材料(實驗組),48 h后掃描電鏡觀察細(xì)胞黏附情況;RT-PCR和Western blot檢測接種于支架的種子細(xì)胞Ⅱ型膠原mRNA和蛋白的表達(dá),并與單純培養(yǎng)的種子細(xì)胞(對照組)進(jìn)行比較。結(jié)果經(jīng)0.5%胰蛋白酶溶液結(jié)構(gòu)重塑后的脫細(xì)胞真皮基質(zhì)支架材料表面光滑、孔隙均勻;孔隙率達(dá)85.4%±2.8%,顯著高于重塑前的支架(72.8%±5.8%)(t=—4.384,P=0.005);細(xì)胞毒性檢測為1級,合格;埋植于大鼠背部皮下后,隨時間延長,組織炎性細(xì)胞數(shù)呈減少趨勢(P0.05)。種子細(xì)胞接種后,掃描電鏡示大量細(xì)胞貼附于支架上,細(xì)胞形態(tài)飽滿,可見表面微絨毛和分泌現(xiàn)象。RT-PCR與Western blot檢測結(jié)果示,實驗組和對照組Ⅱ型膠原mRNA和蛋白表達(dá)量比較,差異均無統(tǒng)計學(xué)意義(t=1.265,P=0.235;t=0.935,P=0.372)。結(jié)論經(jīng)脫細(xì)胞-結(jié)構(gòu)重塑-交聯(lián)-表面修飾制備的脫細(xì)胞真皮基質(zhì)結(jié)構(gòu)良好,種子細(xì)胞能大量黏附于支架材料上,細(xì)胞Ⅱ型膠原表達(dá)水平無明顯改變,有望作為軟骨組織工程支架材料。
[Abstract]:Objective to prepare acellular dermal matrix, detect its cell and histocompatibility, and explore its feasibility as scaffold material for cartilage tissue engineering. The collagen fibers were reconstructed by 0.5% acellular trypsin in 0.5% SDS solution, and the surface was modified by ultrasonic oscillation in 0.5% chondroitin sulfate solution immersed in formaldehyde. Acellular dermal matrix scaffolds were prepared. The changes of porosity before and after structural remodeling were detected by ethanol exclusion and cytotoxicity was detected by MTT method. The materials were implanted subcutaneously into the back of adult Sprague-Dawley rats. To evaluate the histocompatibility, chondrocytes and bone marrow stromal cells of the second passage of New Zealand rabbits co-cultured at the ratio of 3: 7 were used as seed cells. The adhesion of acellular dermal matrix scaffolds was observed by scanning electron microscope 48 hours after implantation. RT-PCR and Western blot were used to detect the expression of type 鈪，

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